Utilization of cathodic hydrogen by hydrogen-oxidizing bacteria

Summary Hydrogen is consumed by methanogenic, sulphate-reducing, and homoacetogenic bacteria and members of these bacterial groups are able to grow chemolithotrophically with hydrogen as sole energy source. Cathodic hydrogen consumption by sulphate-reducing bacteria has been proposed as one of the factors in the anaerobic corrosion of metals. Desulfovibrio spp. were able to utilize cathodic hydrogen from mild steel as the only source of energy for growth with sulphate or nitrate as terminal electron acceptor. Other hydrogen-oxidizing bacteria such as Methanospirillum hungatei, Acetobacterium woodii and Wolinella succinogenes were also able to utilize cathodic hydrogen from mild steel for energy generation and growth. Weight loss studies of mild steel coupons under different growth conditions of Desulfovibrio spp. indicated that hydrogen removal alone is not the cause of corrosion and the depolarization phenomenon probably plays a role only in the initiation of the anaerobic microbial corrosion process.     

Design and optimization of selective serotonin re-uptake inhibitors with high synthetic accessibility Part 1

The reported selective serotonin Re-uptake Inhibitor Litoxetine was used as the starting point in the design of a range of potential SSRIs with high ease of synthetic accessibility. Preparation and subsequent optimization yielded a range of potent and highly selective SSRIs.     

Electron cytochemistry of mycobacteria: Evidence that strongly acidic sulfate groups are present on the surface of H37Rv (virulent) strain ofMycobacterium tuberculosis

Cytochemical characterization of mycobacterial surfaces was carried out on virulent (H37Rv) and avirulent (H37Ra) strains ofMycobacterium tuberculosis. The results were quantified and compared with those obtained with three colony types of the opportunistic pathogenMycobacterium avium. Mycobacterium aurum, a rapidly growing, nonpathogenic species, served as a model for the cytochemical methods. Concanavalin A (ConA) reacted with -d-mannose and -d-glucose residues, whereas negative charged residues were detected with either the ionized ferritin (CF) or the colloidal ferric hydroxide (CIH) method. Strongly acidic sulfate groups were detected by their selective blockage with alcian blue (AB) at pH 1 prior to the CIH labeling at pH 1.8. Weakly acidic groups were demonstrated by AB blockage at pH 2.5 prior to staining with CF stain. Except forM. aurum, all other strains showed a marked heterogeneity in regard to the abundance of their surface labeling. Accessible sulfate groups were present on the cell surface of the virulent H37Rv strain ofM. tuberculosis, but not on the avirulent strain H37Ra. Distribution of ConA receptors, on the other hand, was unrelated to the virulence or pathogenicity of the bacterial strain.     

Variants of the anti-Müllerian hormone gene in a compound heterozygote with the persistent Müllerian duct syndrome and his family

Summary The human genome contains a large number of interspersed simple repeat sequences that are variable in length and can therefore serve as highly informative, polymorphic markers. Typing procedures include conventional multilocus and single locus probing, and polymerase chain reaction aided analysis. We have identified simple sequences in a cosmid clone stemming from the human Y chromosome and consisting of (gata)_n repeats. We have compared these with two equivalent simple repeat loci from chromosome 12. After amplifying the tandemly repeated motifs, we detected between four and eight different alleles at each of the three loci. Codominant inheritance of the alleles was established in family studies and the informativity of the simple repeat loci was determined by typing unrelated individuals. The polymorphisms are suitable for application in linkage studies, practical forensic case work, deficiency cases in paternity determination, and for studying ethnological questions. The mutational mechanisms that bring about changes in simple repeats located both on the autosomes and on the sex chromosomes, are discussed.Professor Dr. Otto Prokop (Humboldt-Universität Berlin) on the occasion of his 70th birthday     

Evidence for pyroglutamyl peptidase I and prolyl endopeptidase activities in the rat insulinoma cell line RINm 5F: lack of relationship with TRH metabolism

Thyrotropin-R eleasing hormone (TRH)-degrading pyroglutamyl peptidase I(PGP I) and prolyl endopeptidase (PE) activities have been demonstrated in rat insulinoma RINm 5F cell line. These two enzymes catalyze the conversion of TRH to Histydyl-Proline-Diketopiperazine and to acid TRH respectively.After cell fractionation, we found all the PGP I and PE activities in the cytosolic fraction. The membranebound PGP II activity is not detectable in the RINm 5F cells. Further investigations on these two cytosolic enzymes show that pyroglutamyl- and proline-containing peptides are inhibitors of each TRH-degrading enzyme.Gelfiltration chromatography on Sephadex G100 shows that PGP I and PE activity have an apparent molecular mass of about 18 kDa and 57 kDa, respectively. Kinetic analysis with TRH as substrate, gives a Km of 44 µM and 235 µM, and a Vmax of 1.49 and 8.80 pmoUmin/µg protein for PGP I and PE, respectively. Immunoreactive TRH, His-Pro-Diketopiperazine and acid TRH levels in the cell line extracts are 2.2 ± 0.9, 22.5 ± 11.1 and 28.7 ± 14.6pg/10⁶ cells, respectively. When cells have been incubated for 2 to 72 hours with a P. E. inhibitor (Z-Gly-Pro-CHN₂) at 5 × 10^–7M, both cell PGP I and PE activities are inhibited. No change in the cellular content of immunoreactive TRH, His-Pro-Diketopiperazine and acid TRH have been observed in treated cells.These data suggest that TRH is not degraded by cytosolic, unspecific PGP I and PE enzymes in RINm 5F. The finding that these cells contain 10 and 13 times more His-Pro-Diketopiperazine and acid TRH than TRH may be an indirect evidence for the existence of another precursor than TRH for these two peptides or of the possibility that TRH can be degraded by other peptidases.Abbreviations TRH Thyrotropin-Releasing Hormone or Thyroliberin - His-Pro-DKP Histidyl-ProlineDiketopiperazine - TRH-OH acid TRH or deamidated TRH - LH-RH Luteinizing Hormone-Releasing Hormone - Z-Gly-Pro-CHN₂ N-benzyloxycarboxyl-Gly-Pro-diazomethylketone - PGP Pyroglutamyl Peptidase, PGP I (EC 3.4.19.3) and PGP II (EC 3.4.19.-) - PE Prolyl Endopeptidase or post-proline cleaving enzyme (EC 3.4.21.26)     

Properties of the complexes of riboflavin 3',5'-bisphosphate and the apoflavodoxins from Megasphaera elsdenii and Desulfovibrio vulgaris

Megasphaera elsdenii and Desulfovibrio vulgaris apoflavodoxins have been reconstituted with riboflavin 3',5'-bisphosphate. Several biochemical and biophysical properties of the complexes have been investigated and the results are compared with the properties of the native proteins. The dissociation constant of the modified complex of M. elsdenii flavodoxin is increased by a factor of about 23 by comparison with that of the native protein. The rate constant for the formation of the complex of M. elsdenii flavodoxin is about 26 times lower than that for the native protein. The redox potential of the transition between the oxidized and semiquinone state is similar to that of the native protein. On the other hand, the redox potential of the semiquinone-hydroquinone transition is about 20 mV more negative than that of the native protein. Absorbance and circular dichroic spectra of the protein-bound artificial prosthetic group and the protein-bound natural prosthetic group are very similar. In both the oxidized and in the fully reduced state only minor differences in interaction between the isoalloxazine ring and the apoprotein for the two flavin derivatives are found by 13C and 15N NMR. 31P-NMR studies show that the 5'-phosphate group of the two flavin derivatives is bound in the same way and that it is dianionic in the complex. In contrast, the 3'-phosphate group in riboflavin 3',5'-bisphosphate is monoanionic or even neutral when bound to the protein. The 3'-phosphate group is also close to or on the surface of the protein. Desulfovibrio vulgaris apoflavodoxin has an affinity for riboflavin 3',5'-bisphosphate which is 10 times higher as compared to Megasphaera elsdenii apoflavodoxin (Ka = 10(8) M-1). Also the association rate constant of Desulfovibrio vulgaris apoprotein and riboflavin 3'5'-bisphosphate is found to be 10 times faster than for the Megasphaera elsdenii flavodoxin reaction. The dissociation behaviour of native Desulfovibrio vulgaris flavodoxin measured under identical conditions as for the riboflavin 3',5'-bisphosphate analog gives a value (Kd approximately equal to 0.2 nM) which is considerably lower than reported earlier [Dubourdieu, M., MacKnight, M. L. & Tollin, G. (1974) Biochem. Biophys. Res. Commun. 60, 649-655]. The results are discussed in the light of the existing crystallographic data of flavodoxins and the recently proposed theory on the regulation of the redox potential in flavoproteins [Moonen, C. T. W., Vervoort, J. & Müller, F. (1984) in Flavins and flavoproteins, pp. 493-496, Walter de Gruyter, Berlin].     

A proton-deuterium exchange study of three types ofDesulfovibrio hydrogenases

Summary Hydrogenases are among the main enzymes involved in bacterial anaerobic corrosion of metals. The study of their mode of action is important for a full comprehension of this phenomenon. The three types ofDesulfovibrio hydrogenases [(Fe), (NiFe), (NiFeSe)] present different patterns in the pH dependence of their activity. The periplasmic enzyme fromDesulfovibrio salexigens and the cytoplasmic enzyme fromDesulfovibrio baculatus both have pH optima at 7.5 for H₂ uptake and 4.0 for H₂ evolution and H⁺–D₂ exchange reaction (measured by membrane-inlet mass-spectrometry). The H₂ to HD ratio at pH above 5.0 is higher than 1.0. The periplasmic hydrogenase fromD. gigas presents the same pH optimum (8.0) for the H⁺–D₂ exchange as for H₂ consumption. In contrast, the enzyme fromD. vulgaris has the highest activity in H₂ production and in the exchange at pH 5.0. Both hydrogenases have a H₂-to-HD ratio below 1.0.     

The relationship between hydrogen metabolism,sulfate reduction and nitrogen fixation in sulfate reducers

Summary Hydrogenase and nitrogenase activities of sulfate-reducing bacteria allow their adaptation to different nutritional habits even under adverse conditions. These exceptional capabilities of adaptation are important factors in the understanding of their predominant role in problems related to anaerobic metal corrosion. Although the D₂–H⁺ exchange reaction indicated thatDesulfovibrio desulfuricans strain Berre-Sol andDesulfovibrio gigas hydrogenases were reversible, the predominant activity in vivo was hydrogen uptake. Hydrogen production was restricted to some particular conditions such as sulfate or nitrogen starvation. Under diazotrophic conditions, a transient hydrogen evolution was followed by uptake when dinitrogen was effectively fixed. In contrast, hydrogen evolution proceeded when acetylene was substituted as the nitrogenase substrate. Hydrogen can thus serve as an electron donor in sulfate reduction and nitrogen metabolism.